Pathlength-independent methods for optically determining material composition

ABSTRACT

A method uses spectroscopy to determine an analyte concentration in a sample. The method includes producing an absorbance spectrum of the sample. The method further includes shifting the absorbance spectrum to zero in a wavelength region. The method further includes subtracting a water or other substance contribution from the absorbance spectrum.

CLAIM OF PRIORITY

[0001] This application claims priority to U.S. Provisional PatentApplication No. 60/341,435 filed Dec. 14, 2001 and U.S. ProvisionalPatent Application No. 60/357,264 filed Feb. 12, 2002, both of which areincorporated in their entireties by reference herein.

BACKGROUND OF THE INVENTION

[0002] 1. Field of the Invention

[0003] The disclosure herein relates generally to methods fordetermining the composition of a material sample by analyzing infraredenergy that has been passed through or emitted from the material sample.

[0004] 2. Description of the Related Art

[0005] When taking measurements of a sample of blood (or othermaterials) in order to assess a particular aspect such as glucosecontent, a number of problems are realized. A sample is collected in/ona test strip, or cuvette, or perhaps by a noninvasive means, whereby asource is applied to the blood sample (or tissue in a noninvasive sense)and the constituents can be analyzed. When the initial readings aretaken, many blood analytes appear in the data and the next step is toidentify which reading correlates to each particular constituent toidentify and quantify the blood analyte of interest.

[0006] For example, a measurement of a blood sample is taken and thefocus is to measure blood glucose in such a way as to identify itscontent and to predict its trend (rising, falling, or sustained). Whenthe sample is measured all sorts of variables influence the data. Toobtain accurate measurements it is helpful to understand whichconstituents are present in the data set, understand their effects onthe analyte that is being measured (such as glucose), and correct forany differences that intrinsic and measuring-device-related variablesmay cause.

SUMMARY OF THE INVENTION

[0007] In certain embodiments, a method uses spectroscopy to determinean analyte concentration in a sample. The method comprises producing anabsorbance spectrum of the sample. The method further comprises shiftingthe absorbance spectrum to zero in a wavelength region. The methodfurther comprises subtracting a water or other substance contributionfrom the absorbance spectrum.

[0008] In certain other embodiments, a method provides pathlengthinsensitive measurements of blood constituents in a sample usinginfrared (IR) spectroscopy. The method comprises providing an absorbancespectrum of the sample. The method further comprises shifting theabsorbance spectrum to zero at an isosbestic wavelength, wherein waterand protein within the sample have approximately equivalent absorptionsat the isosbestic wavelength.

BRIEF DESCRIPTION OF THE DRAWINGS

[0009]FIG. 1 is a schematic view of a noninvasive optical detectionsystem.

[0010]FIG. 2 is a perspective view of a window assembly for use with thenoninvasive detection system.

[0011]FIG. 2A is a plan view of another embodiment of a window assemblyfor use with the noninvasive detection system.

[0012]FIG. 3 is an exploded schematic view of another embodiment of awindow assembly for use with the noninvasive detection system.

[0013]FIG. 4 is a plan view of the window assembly connected to acooling system.

[0014]FIG. 5 is a plan view of the window assembly connected to a coldreservoir.

[0015]FIG. 6 is a cutaway view of a heat sink for use with thenoninvasive detection system.

[0016]FIG. 6A is a cutaway perspective view of a lower portion of thenoninvasive detection system of FIG. 1.

[0017]FIG. 6B is an exploded perspective view of a window mountingsystem for use with the noninvasive optical detection system.

[0018]FIG. 6C is a partial plan view of the window mounting system ofFIG. 6B.

[0019]FIG. 6D is a sectional view of the window mounting system of FIG.6C.

[0020]FIG. 7 is a schematic view of a control system for use with thenoninvasive optical detection system.

[0021]FIG. 8 depicts a first methodology for determining theconcentration of an analyte of interest.

[0022]FIG. 9 depicts a second methodology for determining theconcentration of an analyte of interest.

[0023]FIG. 10 depicts a third methodology for determining theconcentration of an analyte of interest.

[0024]FIG. 11 depicts a fourth methodology for determining theconcentration of an analyte of interest.

[0025]FIG. 12 depicts a fifth methodology for determining theconcentration of an analyte of interest.

[0026]FIG. 13 is a schematic view of a reagentless whole-blood detectionsystem.

[0027]FIG. 14 is a perspective view of one embodiment of a cuvette foruse with the reagentless whole-blood detection system.

[0028]FIG. 15 is a plan view of another embodiment of a cuvette for usewith the reagentless whole-blood detection system.

[0029]FIG. 16 is a disassembled plan view of the cuvette shown in FIG.15.

[0030]FIG. 16A is an exploded perspective view of the cuvette of FIG.15.

[0031]FIG. 17 is a side view of the cuvette of FIG. 15.

[0032]FIG. 18 is a flow diagram of one embodiment of a method for usingspectroscopy to determine an analyte concentration in a sample.

[0033]FIG. 19 is a graph illustrating one isosbestic point betweenapproximately 4.0 and 4.2 μm in the absorbance spectra of water andwhole blood protein.

[0034]FIG. 20 is a graph illustrating another isosbestic point betweenapproximately 9.2 μm and 9.6 μm in the absorbance spectra of water andwhole blood protein.

[0035]FIG. 21 is a graph illustrating progressive removal of free watercontributions from an absorbance spectrum of a sample.

[0036]FIG. 22 is a graph illustrating determination of free protein froman absorbance spectrum of a sample.

[0037]FIG. 23 is a graph illustrating progressive removal of residualinteractive component contributions from an absorbance spectrum of asample.

[0038]FIG. 24 is a graph illustrating an absorbance spectrum withresidual absorbance after glucose spectral data removal used forindividual determination of residual components.

[0039]FIG. 25 is a flow diagram of one embodiment of a method ofproviding pathlength insensitive measurements of blood constituents in asample using infrared (IR) spectroscopy.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

[0040] Although certain preferred embodiments and examples are disclosedbelow, it will be understood by those skilled in the art that theinvention extends beyond the specifically disclosed embodiments to otheralternative embodiments and/or uses of the invention and obviousmodifications and equivalents thereof. Thus, it is intended that thescope of the invention herein disclosed should not be limited by theparticular disclosed embodiments described below.

[0041] Embodiments are described herein using flow diagrams that havesteps in a particular order, and the order of the steps in the flowdiagrams are not to be considered to be limiting. Other methods withdifferent orders of steps are also compatible with embodiments describedherein. In addition, other methods with additional steps are alsocompatible with embodiments described herein.

I. Overview of Analyte Detection Systems

[0042] Disclosed herein are analyte detection systems, including anoninvasive system discussed largely in part A below and a whole-bloodsystem discussed largely in part B below. Also disclosed are variousmethods, including methods for detecting the concentration of an analytein a material sample. Both the noninvasive system/method and thewhole-blood system/method can employ optical measurement. As used hereinwith reference to measurement apparatus and methods, “optical” is abroad term and is used in its ordinary sense and refers, withoutlimitation, to identification of the presence or concentration of ananalyte in a material sample without requiring a chemical reaction totake place. As discussed in more detail below, the two approaches eachcan operate independently to perform an optical analysis of a materialsample. The two approaches can also be combined in an apparatus, or thetwo approaches can be used together to perform different steps of amethod.

[0043] In one embodiment, the two approaches are combined to performcalibration of an apparatus, e.g., of an apparatus that employs anoninvasive approach. In another embodiment, an advantageous combinationof the two approaches performs an invasive measurement to achievegreater accuracy and a whole-blood measurement to minimize discomfort tothe patient. For example, the whole-blood technique may be more accuratethan the noninvasive technique at certain times of the day, e.g., atcertain times after a meal has been consumed, or after a drug has beenadministered.

[0044] It should be understood, however, that any of the discloseddevices may be operated in accordance with any suitable detectionmethodology, and that any disclosed method may be employed in theoperation of any suitable device. Furthermore, the disclosed devices andmethods are applicable in a wide variety of situations or modes ofoperation, including but not limited to invasive, noninvasive,intermittent or continuous measurement, subcutaneous implantation,wearable detection systems, or any combination thereof.

[0045] Any method which is described and illustrated herein is notlimited to the exact sequence of acts described, nor is it necessarilylimited to the practice of all of the acts set forth. Other sequences ofevents or acts, or less than all of the events, or simultaneousoccurrence of the events, may be utilized in practicing the method(s) inquestion.

[0046] A. Noninvasive System

[0047] 1. Monitor Structure

[0048]FIG. 1 depicts a noninvasive optical detection system (hereinafter“noninvasive system”) 10 in a presently preferred configuration. Thedepicted noninvasive system 10 is particularly suited for noninvasivelydetecting the concentration of an analyte in a material sample S, byobserving the infrared energy emitted by the sample, as will bediscussed in further detail below.

[0049] As used herein, the term “noninvasive” is a broad term and isused in its ordinary sense and refers, without limitation, to analytedetection devices and methods which have the capability to determine theconcentration of an analyte in in-vivo tissue samples or bodily fluids.It should be understood, however, that the noninvasive system 10disclosed herein is not limited to noninvasive use, as the noninvasivesystem 10 may be employed to analyze an in-vitro fluid or tissue samplewhich has been obtained invasively or noninvasively. As used herein, theterm “invasive” (or, alternatively, “traditional”) is a broad term andis used in its ordinary sense and refers, without limitation, to analytedetection methods which involve the removal of fluid samples through theskin. As used herein, the term “material sample” is a broad term and isused in its ordinary sense and refers, without limitation, to anycollection of material which is suitable for analysis by the noninvasivesystem 10. For example, the material sample S may comprise a tissuesample, such as a human forearm, placed against the noninvasive system10. The material sample S may also comprise a volume of a bodily fluid,such as whole blood, blood component(s), interstitial fluid orintercellular fluid obtained invasively, or saliva or urine obtainednoninvasively, or any collection of organic or inorganic material. Asused herein, the term “analyte” is a broad term and is used in itsordinary sense and refers, without limitation, to any chemical speciesthe presence or concentration of which is sought in the material sampleS by the noninvasive system 10. For example, the analyte(s) which may bedetected by the noninvasive system 10 include but not are limited toglucose, ethanol, insulin, water, carbon dioxide, blood oxygen,cholesterol, bilirubin, ketones, fatty acids, lipoproteins, albumin,urea, creatinine, white blood cells, red blood cells, hemoglobin,oxygenated hemoglobin, carboxyhemoglobin, organic molecules, inorganicmolecules, pharmaceuticals, cytochrome, various proteins andchromophores, microcalcifications, electrolytes, sodium, potassium,chloride, bicarbonate, and hormones. As used herein to describemeasurement techniques, the term “continuous” is a broad term and isused in its ordinary sense and refers, without limitation, to the takingof discrete measurements more frequently than about once every 10minutes, and/or the taking of a stream or series of measurements orother data over any suitable time interval, for example, over aninterval of one to several seconds, minutes, hours, days, or longer. Asused herein to describe measurement techniques, the term “intermittent”is a broad term and is used in its ordinary sense and refers, withoutlimitation, to the taking of measurements less frequently than aboutonce every 10 minutes.

[0050] The noninvasive system 10 preferably comprises a window assembly12, although in some embodiments the window assembly 12 may be omitted.One function of the window assembly 12 is to permit infrared energy E toenter the noninvasive system 10 from the sample S when it is placedagainst an upper surface 12 a of the window assembly 12. The windowassembly 12 includes a heater layer (see discussion below) which isemployed to heat the material sample S and stimulate emission ofinfrared energy therefrom. A cooling system 14, preferably comprising aPeltier-type thermoelectric device, is in thermally conductive relationto the window assembly 12 so that the temperature of the window assembly12 and the material sample S can be manipulated in accordance with adetection methodology discussed in greater detail below. The coolingsystem 14 includes a cold surface 14 a which is in thermally conductiverelation to a cold reservoir 16 and the window assembly 12, and a hotsurface 14 b which is in thermally conductive relation to a heat sink18.

[0051] As the infrared energy E enters the noninvasive system 10, itfirst passes through the window assembly 12, then through an opticalmixer 20, and then through a collimator 22. The optical mixer 20preferably comprises a light pipe having highly reflective innersurfaces which randomize the directionality of the infrared energy E asit passes therethrough and reflects against the mixer walls. Thecollimator 22 also comprises a light pipe having highly-reflective innerwalls, but the walls diverge as they extend away from the mixer 20. Thedivergent walls cause the infrared energy E to tend to straighten as itadvances toward the wider end of the collimator 22, due to the angle ofincidence of the infrared energy when reflecting against the collimatorwalls.

[0052] From the collimator 22 the infrared energy E passes through anarray of filters 24, each of which allows only a selected wavelength orband of wavelengths to pass therethrough. These wavelengths/bands areselected to highlight or isolate the absorptive effects of the analyteof interest in the detection methodology discussed in greater detailbelow. Each filter 24 is preferably in optical communication with aconcentrator 26 and an infrared detector 28. The concentrators 26 havehighly reflective, converging inner walls which concentrate the infraredenergy as it advances toward the detectors 28, increasing the density ofthe energy incident upon the detectors 28.

[0053] The detectors 28 are in electrical communication with a controlsystem 30 which receives electrical signals from the detectors 28 andcomputes the concentration of the analyte in the sample S. The controlsystem 30 is also in electrical communication with the window 12 andcooling system 14, so as to monitor the temperature of the window 12and/or cooling system 14 and control the delivery of electrical power tothe window 12 and cooling system 14.

[0054] a. Window Assembly

[0055] A preferred configuration of the window assembly 12 is shown inperspective, as viewed from its underside (in other words, the side ofthe window assembly 12 opposite the sample S), in FIG. 2. The windowassembly 12 generally comprises a main layer 32 formed of a highlyinfrared-transmissive material and a heater layer 34 affixed to theunderside of the main layer 32. The main layer 32 is preferably formedfrom diamond, most preferably from chemical-vapor-deposited (“CVD”)diamond, with a preferred thickness of about 0.25 millimeters. In otherembodiments alternative materials which are highlyinfrared-transmissive, such as silicon or germanium, may be used informing the main layer 32.

[0056] The heater layer 34 preferably comprises bus bars 36 located atopposing ends of an array of heater elements 38. The bus bars 36 are inelectrical communication with the elements 38 so that, upon connectionof the bus bars 36 to a suitable electrical power source (not shown) acurrent may be passed through the elements 38 to generate heat in thewindow assembly 12. The heater layer 34 may also include one or moretemperature sensors (not shown), such as thermistors or resistancetemperature devices (RTDs), to measure the temperature of the windowassembly 12 and provide temperature feedback to the control system 30(see FIG. 1).

[0057] Still referring to FIG. 2, the heater layer 34 preferablycomprises a first adhesion layer of gold or platinum (hereinafterreferred to as the “gold” layer) deposited over an alloy layer which isapplied to the main layer 32. The alloy layer comprises a materialsuitable for implementation of the heater layer 34, such as, by way ofexample, 10/90 titanium/tungsten, titanium/platinum, nickel/chromium, orother similar material. The gold layer preferably has a thickness ofabout 4000 Å, and the alloy layer preferably has a thickness rangingbetween about 300 Å and about 500 Å. The gold layer and/or the alloylayer may be deposited onto the main layer 32 by chemical depositionincluding, but not necessarily limited to, vapor deposition, liquiddeposition, plating, laminating, casting, sintering, or other forming ordeposition methodologies well known to those or ordinary skill in theart. If desired, the heater layer 34 may be covered with an electricallyinsulating coating which also enhances adhesion to the main layer 32.One preferred coating material is aluminum oxide. Other acceptablematerials include, but are not limited to, titanium dioxide or zincselenide.

[0058] The heater layer 34 may incorporate a variable pitch distancebetween centerlines of adjacent heater elements 38 to maintain aconstant power density, and promote a uniform temperature, across theentire layer 34. Where a constant pitch distance is employed, thepreferred distance is at least about 50-100 microns. Although the heaterelements 38 generally have a preferred width of about 25 microns, theirwidth may also be varied as needed for the same reasons stated above.

[0059] Alternative structures suitable for use as the heater layer 34include, but are not limited to, thermoelectric heaters, radiofrequency(RF) heaters, infrared radiation heaters, optical heaters, heatexchangers, electrical resistance heating grids, wire bridge heatinggrids, or laser heaters. Whichever type of heater layer is employed, itis preferred that the heater layer obscures about 10% or less of thewindow assembly 12.

[0060] In a preferred embodiment, the window assembly 12 comprisessubstantially only the main layer 32 and the heater layer 34. Thus, wheninstalled in an optical detection system such as the noninvasive system10 shown in FIG. 1, the window assembly 12 will facilitate a minimallyobstructed optical path between a (preferably flat) upper surface 12 aof the window assembly 12 and the infrared detectors 28 of thenoninvasive system 10. The optical path 32 in the preferred noninvasivesystem 10 proceeds only through the main layer 32 and heater layer 34 ofthe window assembly 12 (including any antireflective, index-matching,electrical insulating or protective coatings applied thereto or placedtherein), through the optical mixer 20 and collimator 22 and to thedetectors 28.

[0061]FIG. 2A shows another embodiment of the window assembly 12, thatmay be used in place of the window assembly 12 depicted in FIG. 2. Thewindow assembly 12 shown in FIG. 2A may be similar to that shown in FIG.2, except as described below. In the embodiment of FIG. 2A the mainlayer 32 has a preferred thickness of up to about 0.012″ and morepreferably about 0.010″ or less. The heater layer 34 may also includeone or more resistance temperature devices (RTD's) 55 to measure thetemperature of the window assembly 12 and provide temperature feedbackto a control system 30. The RTDs 55 terminate in RTD connection pads 57.

[0062] In the embodiment of FIG. 2A, the heater elements 38 aretypically provided with a width of about 25 microns. The pitch distanceseparating centerlines of adjacent heater elements 38 may be reduced,and/or the width of the heater elements 38 may be increased, in theregions of the window assembly 12 near the point(s) of contact with thethermal diffuser 410 (see FIGS. 6B-6D and discussion below). Thisarrangement advantageously promotes an isothermal temperature profile atthe upper surface of the main layer 32 despite thermal contact with thethermal diffuser.

[0063] The embodiment shown in FIG. 2A includes a plurality of heaterelements 38 of substantially equal width which are variably spacedacross the width of the main layer 32. In the embodiment of FIG. 2A, thecenterlines of the heater elements 38 are spaced at a first pitchdistance of about 0.0070″ at peripheral portions 34 a of the heaterlayer 34, and at a second pitch distance of about 0.015″ at a centralportion 34 b of the main layer 32. The heater elements 38 closest to thecenter are preferably sufficiently spaced to allow the RTDs 55 to extendtherebetween. In the embodiment of FIG. 2A, the main layer 32 includesperipheral regions 32 a which extend about 0.053″ from the outermostheater element on each side of the heater layer 34 to the adjacent edgeof the main layer 32. As shown, the bus bars 36 are preferablyconfigured and segmented to allow space for the RTDs 55 and the RTDconnection pads 57, in intermediate gaps 36 a. The RTDs 55 preferablyextend into the array of heater elements 38 by distance that is slightlylonger than half of the length of an individual heater element 38. Inalternative embodiments, the RTDs 55 may be located at the edges of themain layer 32, or at other locations as desired for a particularnoninvasive system.

[0064] With continued reference to FIG. 2A, the peripheral regions ofthe main layer 32 may include metallized edge portions 35 forfacilitating connection to the diffuser 410 (discussed below inconnection with FIGS. 6B-6D). The metallized edge portions 35 may beformed by the same or similar processes used in forming the heaterelements 38 and RTDs 55. In the embodiment of FIG. 2A, the edge portions35 are typically between about 0.040″ and about 0.060″ wide by about0.450″ and about 0.650″ long, and in one embodiment, they are about0.050″ by about 0.550″. Other dimensions may be appropriately used solong as the window assembly 12 may be joined in thermal communicationwith the diffuser 410 as needed.

[0065] In the embodiment shown in FIG. 2A, the main layer 32 is about0.690″ long by about 0.571″ wide, and the heater layer (excluding themetallized edge portions 35) is about 0.640″ long by about 0.465″ wide.The main layer 32 is about 0.010″-0.012″ thick, and is advantageouslythinner than about 0.010″ where possible. Each heater element 38 isabout 0.570″ long, and each peripheral region 34 a is about 0.280″ wide.These dimensions are merely exemplary; of course, other dimensions maybe used as desired.

[0066]FIG. 3 depicts an exploded side view of an alternativeconfiguration for the window assembly 12, which may be used in place ofthe configuration shown in FIG. 2. The window assembly 12 depicted inFIG. 3 includes near its upper surface (the surface intended for contactwith the sample S) a highly infrared-transmissive, thermally conductivespreader layer 42. Underlying the spreader layer 42 is a heater layer44. A thin electrically insulating layer (not shown), such as layer ofaluminum oxide, titanium dioxide or zinc selenide, may be disposedbetween the heater layer 44 and the spreader layer 42. (An aluminumoxide layer also increases adhesion of the heater layer 44 to thespreader layer 42.) Adjacent to the heater layer 44 is a thermalinsulating and impedance matching layer 46. Adjacent to the thermalinsulating layer 46 is a thermally conductive inner layer 48. Thespreader layer 42 is coated on its top surface with a thin layer ofprotective coating 50. The bottom surface of the inner layer 48 iscoated with a thin overcoat layer 52. Preferably, the protective coating50 and the overcoat layer 52 have antireflective properties.

[0067] The spreader layer 42 is preferably formed of a highlyinfrared-transmissive material having a high thermal conductivitysufficient to facilitate heat transfer from the heater layer 44uniformly into the material sample S when it is placed against thewindow assembly 12. Other effective materials include, but are notlimited to, CVD diamond, diamondlike carbon, gallium arsenide,germanium, and other infrared-transmissive materials having sufficientlyhigh thermal conductivity. Preferred dimensions for the spreader layer42 are about one inch in diameter and about 0.010 inch thick. As shownin FIG. 3, a preferred embodiment of the spreader layer 42 incorporatesa beveled edge. Although not required, an approximate 45-degree bevel ispreferred.

[0068] The protective layer 50 is intended to protect the top surface ofthe spreader layer 42 from damage. Ideally, the protective layer ishighly infrared-transmissive and highly resistant to mechanical damage,such as scratching or abrasion. It is also preferred that the protectivelayer 50 and the overcoat layer 52 have high thermal conductivity andantireflective and/or index-matching properties. A satisfactory materialfor use as the protective layer 50 and the overcoat layer 52 is themulti-layer Broad Band Anti-Reflective Coating produced by DepositionResearch Laboratories, Inc. of St. Charles, Miss. Diamondlike carboncoatings are also suitable.

[0069] Except as noted below, the heater layer 44 is generally similarto the heater layer 34 employed in the window assembly shown in FIG. 2.Alternatively, the heater layer 44 may comprise a dopedinfrared-transmissive material, such as a doped silicon layer, withregions of higher and lower resistivity. The heater layer 44 preferablyhas a resistance of about 2 ohms and has a preferred thickness of about1,500 angstroms. A preferred material for forming the heater layer 44 isa gold alloy, but other acceptable materials include, but are notlimited to, platinum, titanium, tungsten, copper, and nickel.

[0070] The thermal insulating layer 46 prevents the dissipation of heatfrom the heater element 44 while allowing the cooling system 14 toeffectively cool the material sample S (see FIG. 1). This layer 46comprises a material having thermally insulative (e.g., lower thermalconductivity than the spreader layer 42) and infrared transmissivequalities. A preferred material is a germanium-arsenic-selenium compoundof the calcogenide glass family known as AMTIR-1 produced by AmorphousMaterials, Inc. of Garland, Texas. The pictured embodiment has adiameter of about 0.85 inches and a preferred thickness in the range ofabout 0.005 to about 0.010 inches. As heat generated by the heater layer44 passes through the spreader layer 42 into the material sample S, thethermal insulating layer 46 insulates this heat.

[0071] The inner layer 48 is formed of thermally conductive material,preferably crystalline silicon formed using a conventional floatzonecrystal growth method. The purpose of the inner layer 48 is to serve asa cold-conducting mechanical base for the entire layered windowassembly.

[0072] The overall optical transmission of the window assembly 12 shownin FIG. 3 is preferably at least 70%. The window assembly 12 of FIG. 3is preferably held together and secured to the noninvasive system 10 bya holding bracket (not shown). The bracket is preferably formed of aglass-filled plastic, for example Ultem 2300, manufactured by GeneralElectric. Ultem 2300 has low thermal conductivity which prevents heattransfer from the layered window assembly 12.

[0073] b. Cooling System

[0074] The cooling system 14 (see FIG. 1) preferably comprises aPeltier-type thermoelectric device. Thus, the application of anelectrical current to the preferred cooling system 14 causes the coldsurface 14 a to cool and causes the opposing hot surface 14 b to heatup. The cooling system 14 cools the window assembly 12 via the situationof the window assembly 12 in thermally conductive relation to the coldsurface 14 a of the cooling system 14. It is contemplated that thecooling system 14, the heater layer 34, or both, can be operated toinduce a desired time-varying temperature in the window assembly 12 tocreate an oscillating thermal gradient in the sample S, in accordancewith various analyte-detection methodologies discussed herein.

[0075] Preferably, the cold reservoir 16 is positioned between thecooling system 14 and the window assembly 12, and functions as a thermalconductor between the system 14 and the window assembly 12. The coldreservoir 16 is formed from a suitable thermally conductive material,preferably brass. Alternatively, the window assembly 12 can be situatedin direct contact with the cold surface 14 a of the cooling system 14.

[0076] In alternative embodiments, the cooling system 14 may comprise aheat exchanger through which a coolant, such as air, nitrogen or chilledwater, is pumped, or a passive conduction cooler such as a heat sink. Asa further alternative, a gas coolant such as nitrogen may be circulatedthrough the interior of the noninvasive system 10 so as to contact theunderside of the window assembly 12 (see FIG. 1) and conduct heattherefrom.

[0077]FIG. 4 is a top schematic view of a preferred arrangement of thewindow assembly 12 (of the types shown in FIG. 2 or 2A) and the coldreservoir 16, and FIG. 5 is a top schematic view of an alternativearrangement in which the window assembly 12 directly contacts thecooling system 14. The cold reservoir 16/cooling system 14 preferablycontacts the underside of the window assembly 12 along opposing edgesthereof, on either side of the heater layer 34. With thermalconductivity thus established between the window assembly 12 and thecooling system 14, the window assembly can be cooled as needed duringoperation of the noninvasive system 10. In order to promote asubstantially uniform or isothermal temperature profile over the uppersurface of the window assembly 12, the pitch distance betweencenterlines of adjacent heater elements 38 may be made smaller (therebyincreasing the density of heater elements 38) near the region(s) ofcontact between the window assembly 12 and the cold reservoir 16/coolingsystem 14. As a supplement or alternative, the heater elements 38themselves may be made wider near these regions of contact. As usedherein, “isothermal” is a broad term and is used in its ordinary senseand refers, without limitation, to a condition in which, at a givenpoint in time, the temperature of the window assembly 12 or otherstructure is substantially uniform across a surface intended forplacement in thermally conductive relation to the material sample S.Thus, although the temperature of the structure or surface may fluctuateover time, at any given point in time the structure or surface maynonetheless be isothermal.

[0078] The heat sink 18 drains waste heat from the hot surface 14 b ofthe cooling system 16 and stabilizes the operational temperature of thenoninvasive system 10. The preferred heat sink 18 (see FIG. 6) comprisesa hollow structure formed from brass or any other suitable materialhaving a relatively high specific heat and high heat conductivity. Theheat sink 18 has a conduction surface 18 a which, when the heat sink 18is installed in the noninvasive system 18, is in thermally conductiverelation to the hot surface 14 b of the cooling system 14 (see FIG. 1).A cavity 54 is formed in the heat sink 18 and preferably contains aphase-change material (not shown) to increase the capacity of the sink18. A preferred phase change material is a hydrated salt, such ascalciumchloride hexahydrate, available under the name TH29 from PCMThermal Solutions, Inc., of Naperville, Ill. Alternatively, the cavity54 may be omitted to create a heat sink 18 comprising a solid, unitarymass. The heat sink 18 also forms a number of fins 56 to furtherincrease the conduction of heat from the sink 18 to surrounding air.

[0079] Alternatively, the heat sink 18 may be formed integrally with theoptical mixer 20 and/or the collimator 22 as a unitary mass of rigid,heat-conductive material such as brass or aluminum. In such a heat sink,the mixer 20 and/or collimator 22 extend axially through the heat sink18, and the heat sink defines the inner walls of the mixer 20 and/orcollimator 22. These inner walls are coated and/or polished to haveappropriate reflectivity and nonabsorbance in infrared wavelengths aswill be further described below. Where such a unitary heatsink-mixer-collimator is employed, it is desirable to thermally insulatethe detector array from the heat sink.

[0080] It should be understood that any suitable structure may beemployed to heat and/or cool the material sample S, instead of or inaddition to the window assembly 12/cooling system 14 disclosed above, solong a proper degree of cycled heating and/or cooling are imparted tothe material sample S. In addition other forms of energy, such as butnot limited to light, radiation, chemically induced heat, friction andvibration, may be employed to heat the material sample S. It will befurther appreciated that heating of the sample can achieved by anysuitable method, such as convection, conduction, radiation, etc.

[0081] c. Window Mounting System

[0082]FIG. 6B illustrates an exploded view of a window mounting system400 which, in one embodiment, is employed as part of the noninvasivesystem 10 disclosed above. Where employed in connection with thenoninvasive system 10, the window mounting system 400 supplements or,where appropriate, replaces any of the window assembly 12, coolingsystem 14, cold reservoir 16 and heat sink 18 shown in FIG. 1. In oneembodiment, the window mounting system 400 is employed in conjunctionwith the window assembly 12 depicted in FIG. 2A; in alternativeembodiments, the window assemblies shown in FIGS. 2 and 3 and describedabove may also be used in conjunction with the window mounting system400 illustrated in FIG. 6B.

[0083] In the window mounting system 400, the window assembly 12 isphysically and electrically connected (typically by soldering) to afirst printed circuit board (“first PCB”) 402. The window assembly 12 isalso in thermally conductive relation (typically by contact) to athermal diffuser 410. The window assembly may also be fixed to thediffuser 410 by soldering.

[0084] The thermal diffuser 410 generally comprises a heat spreaderlayer 412 which, as mentioned, preferably contacts the window assembly12, and a conductive layer 414 which is typically soldered to the heatspreader layer 412. The conductive layer 414 may then be placed indirect contact with a cold side 418 a of a thermoelectric cooler (TEC)418 or other cooling device. The TEC 418, which in one embodimentcomprises a 25 W TEC manufactured by MELCOR, is in electricalcommunication with a second PCB 403, which includes TEC power leads 409and TEC power terminals 411 for connection of the TEC 418 to anappropriate power source (not shown). The second PCB 403 also includescontacts 408 for connection with RTD terminals 407 (see FIG. 6C) of thefirst PCB 402. A heat sink 419, which may take the form of theillustrated water jacket, the heat sink 18 shown in FIG. 6, any otherheat sink structures mentioned herein, or any other appropriate device,is in thermal communication with a hot side 418 b of the TEC 418 (orother cooling device), in order to remove any excess heat created by theTEC 418.

[0085]FIG. 6C illustrates a plan view of the interconnection of thewindow assembly 12, the first PCB 402, the diffuser 410 and thethermoelectric cooler 418. The first PCB includes RTD bonding leads 406and heater bonding pads 404 which permit attachment of the RTDs 55 andbus bars 36, respectively, of the window assembly 12 to the first PCB402 via soldering or other conventional techniques. Electricalcommunication is thus established between the heater elements 38 of theheater layer 34, and heater terminals 405 formed in the heater bondingpads 404. Similarly, electrical communication is established between theRTDs 55 and RTD terminals 407 formed at the ends of the RTD bondingleads 406. Electrical connections can be established with the heaterelements 38 and the RTDs 55 via simple connection to the terminals 405,407 of the first PCB 402.

[0086] With further reference to FIGS. 2A and 6B-6C, the heat spreaderlayer 412 of the thermal diffuser 410 contacts the underside of the mainlayer 32 of the window assembly 12 via a pair of rails 416. The rails416 may contact the main layer 32 at the metallized edge portions 35, orat any other appropriate location. The physical and thermal connectionbetween the rails 416 and the window main layer 32 may be achieved bysoldering, as indicated above. Alternatively, the connection may beachieved by an adhesive such as epoxy, or any other appropriate method.The material chosen for the window main layer 32 is preferablysufficiently thermally conductive that heat may be quickly removed fromthe main layer 32 through the rails 416, the diffuser 410, and the TEC128.

[0087]FIG. 6D shows a cross-sectional view of the assembly of FIG. 6Cthrough line 22-22. As can be seen in FIG. 6D, the window assembly 12contacts the rails 416 of the heat spreader layer 412. The conductivelayer 414 underlies the spreader layer 412 and may comprise protrusions426 configured to extend through openings 424 formed in the spreaderlayer 412. The openings 424 and protrusions 426 are sized to leavesufficient expansion space therebetween, to allow expansion andcontraction of the conductive layer 414 without interference with, orcausing deformation of, the window assembly 12 or the heat spreaderlayer 412. Moreover, the protrusions 426 and openings 424 coact toprevent displacement of the spreader layer 412 with respect to theconductive layer 414 as the conductive layer 414 expands and contracts.

[0088] The thermal diffuser 410 provides a thermal impedance between theTEC 418 and the window assembly 12, which impedance is selected to drainheat from the window assembly at a rate proportional to the power outputof the heater layer 34. In this way, the temperature of the main layer32 can be rapidly cycled between a “hot” and a “cold” temperatures,thereby allowing a time-varying thermal gradient to be induced in asample S placed against the window assembly 12.

[0089] The heat spreader layer 412 is preferably made of a materialwhich has substantially the same coefficient of thermal expansion as thematerial used to form the window assembly main layer 32, within theexpected operating temperature range. Preferably, both the material usedto form the main layer 32 and the material used to form the heatspreader layer 412 have substantially the same, extremely low,coefficient of thermal expansion. For this reason, CVD diamond ispreferred for the main layer 32 (as mentioned above); with a CVD diamondmain layer 32 the preferred material for the heat spreader layer 412 isInvar. Invar advantageously has an extremely low coefficient of thermalexpansion and a relatively high thermal conductivity. Because Invar is ametal, the main layer 32 and the heat spreader layer 412 can bethermally bonded to one another with little difficulty. Alternatively,other materials may be used for the heat spreader layer 412; forexample, any of a number of glass and ceramic materials with lowcoefficients of thermal expansion may be employed.

[0090] The conductive layer 414 of the thermal diffuser 410 is typicallya highly thermally conductive material such as copper (or,alternatively, other metals or non-metals exhibiting comparable thermalconductivities). The conductive layer 414 is typically soldered orotherwise bonded to the underside of the heat spreader layer 412.

[0091] In the illustrated embodiment, the heat spreader layer 412 may beconstructed according to the following dimensions, which are to beunderstood as exemplary; accordingly the dimensions may be varied asdesired. The heat spreader layer 412 has an overall length and width ofabout 1.170″, with a central opening of about 0.590″ long by 0.470″wide. Generally, the heat spreader layer 412 is about 0.030″ thick;however, the rails 416 extend a further 0.045″ above the basic thicknessof the heat spreader layer 412. Each rail 416 has an overall length ofabout 0.710″; over the central 0.525″ of this length each rail 416 isabout 0.053″ wide. On either side of the central width each rail 416tapers, at a radius of about 0.6″, down to a width of about 0.023″. Eachopening 424 is about 0.360″ long by about 0.085″ wide, with cornersrounded at a radius of about 0.033″.

[0092] In the illustrated embodiment, conductive layer 414 may beconstructed according to the following dimensions, which are to beunderstood as exemplary; accordingly the dimensions may be varied asdesired. The conductive layer 414 has an overall length and width ofabout 1.170″, with a central opening of about 0.590″ long by 0.470″wide. Generally, the conductive layer 412 is about 0.035″ thick;however, the protrusions 426 extend a further 0.075″-0.085″ above thebasic thickness of the conductive layer 414. Each protrusion 426 isabout 0.343″ long by about 0.076″ wide, with corners rounded at a radiusof about 0.035″.

[0093] As shown in FIG. 6B, first and second clamping plates 450 and 452may be used to clamp the portions of the window mounting system 400 toone another. For example, the second clamping plate 452 is configured toclamp the window assembly 12 and the first PCB 402 to the diffuser 410with screws or other fasteners extending through the openings shown inthe second clamping plate 452, the heat spreader layer 412 and theconductive layer 414. Similarly, the first clamping plate 450 isconfigured overlie the second clamping plate 452 and clamp the rest ofthe window mounting system 400 to the heat sink 419, thus sandwichingthe second clamping plate 452, the window assembly 12, the first PCB402, the diffuser 410, the second PCB 403, and the TEC 418 therebetween.The first clamping plate 450 prevents undesired contact between thesample S and any portion of the window mounting system 400, other thanthe window assembly 12 itself. Other mounting plates and mechanisms mayalso be used as desired.

[0094] d. Optics

[0095] As shown in FIG. 1, the optical mixer 20 comprises a light pipewith an inner surface coating which is highly reflective and minimallyabsorptive in infrared wavelengths, preferably a polished gold coating,although other suitable coatings may be used where other wavelengths ofelectromagnetic radiation are employed. The pipe itself may befabricated from a another rigid material such as aluminum or stainlesssteel, as long as the inner surfaces are coated or otherwise treated tobe highly reflective. Preferably, the optical mixer 20 has a rectangularcross-section (as taken orthogonal to the longitudinal axis A-A of themixer 20 and the collimator 22), although other cross-sectional shapes,such as other polygonal shapes or circular or elliptical shapes, may beemployed in alternative embodiments. The inner walls of the opticalmixer 20 are substantially parallel to the longitudinal axis A-A of themixer 20 and the collimator 22. The highly reflective and substantiallyparallel inner walls of the mixer 20 maximize the number of times theinfrared energy E will be reflected between the walls of the mixer 20,thoroughly mixing the infrared energy E as it propagates through themixer 20. In a presently preferred embodiment, the mixer 20 is about 1.2inches to 2.4 inches in length and its cross-section is a rectangle ofabout 0.4 inches by about 0.6 inches. Of course, other dimensions may beemployed in constructing the mixer 20. In particular it is beadvantageous to miniaturize the mixer or otherwise make it as small aspossible

[0096] Still referring to FIG. 1, the collimator 22 comprises a tubewith an inner surface coating which is highly reflective and minimallyabsorptive in infrared wavelengths, preferably a polished gold coating.The tube itself may be fabricated from a another rigid material such asaluminum, nickel or stainless steel, as long as the inner surfaces arecoated or otherwise treated to be highly reflective. Preferably, thecollimator 22 has a rectangular cross-section, although othercross-sectional shapes, such as other polygonal shapes or circular,parabolic or elliptical shapes, may be employed in alternativeembodiments. The inner walls of the collimator 22 diverge as they extendaway from the mixer 20. Preferably, the inner walls of the collimator 22are substantially straight and form an angle of about 7 degrees withrespect to the longitudinal axis A-A. The collimator 22 aligns theinfrared energy E to propagate in a direction that is generally parallelto the longitudinal axis A-A of the mixer 20 and the collimator 22, sothat the infrared energy E will strike the surface of the filters 24 atan angle as close to 90 degrees as possible.

[0097] In a presently preferred embodiment, the collimator is about 7.5inches in length. At its narrow end 22 a, the cross-section of thecollimator 22 is a rectangle of about 0.4 inches by 0.6 inches. At itswide end 22 b, the collimator 22 has a rectangular cross-section ofabout 1.8 inches by 2.6 inches. Preferably, the collimator 22 aligns theinfrared energy E to an angle of incidence (with respect to thelongitudinal axis A-A) of about 0-15 degrees before the energy Eimpinges upon the filters 24. Of course, other dimensions or incidenceangles may be employed in constructing and operating the collimator 22.

[0098] With further reference to FIGS. 1 and 6A, each concentrator 26comprises a tapered surface oriented such that its wide end 26 a isadapted to receive the infrared energy exiting the corresponding filter24, and such that its narrow end 26 b is adjacent to the correspondingdetector 28. The inward-facing surfaces of the concentrators 26 have aninner surface coating which is highly reflective and minimallyabsorptive in infrared wavelengths, preferably a polished gold coating.The concentrators 26 themselves may be fabricated from a another rigidmaterial such as aluminum, nickel or stainless steel, so long as theirinner surfaces are coated or otherwise treated to be highly reflective.

[0099] Preferably, the concentrators 26 have a rectangular cross-section(as taken orthogonal to the longitudinal axis A-A), although othercross-sectional shapes, such as other polygonal shapes or circular,parabolic or elliptical shapes, may be employed in alternativeembodiments. The inner walls of the concentrators converge as theyextend toward the narrow end 26 b. Preferably, the inner walls of thecollimators 26 are substantially straight and form an angle of about 8degrees with respect to the longitudinal axis A-A. Such a configurationis adapted to concentrate infrared energy as it passes through theconcentrators 26 from the wide end 26 a to the narrow end 26 b, beforereaching the detectors 28.

[0100] In a presently preferred embodiment, each concentrator 26 isabout 1.5 inches in length. At the wide end 26 a, the cross-section ofeach concentrator 26 is a rectangle of about 0.6 inches by 0.57 inches.At the narrow end 26 b, each concentrator 26 has a rectangularcross-section of about 0.177 inches by 0.177 inches. Of course, otherdimensions or incidence angles may be employed in constructing theconcentrators 26.

[0101] e. Filters

[0102] The filters 24 preferably comprise standard interference-typeinfrared filters, widely available from manufacturers such as OpticalCoating Laboratory, Inc. (“OCLI”) of Santa Rosa, Calif. In theembodiment illustrated in FIG. 1, a 3×4 array of filters 24 ispositioned above a 3×4 array of detectors 28 and concentrators 26. Asemployed in this embodiment, the filters 24 are arranged in four groupsof three filters having the same wavelength sensitivity. These fourgroups have bandpass center wavelengths of 7.15 μm±0.03 μm, 8.40 μm±0.03μm, 9.48 μm±0.04 μm, and 11.10 μm±0.04 μm, respectively, whichcorrespond to wavelengths around which water and glucose absorbelectromagnetic radiation. Typical bandwidths for these filters rangefrom 0.20 μm to 0.50 μm.

[0103] In an alternative embodiment, the array of wavelength-specificfilters 24 may be replaced with a single Fabry-Perot interferometer,which can provide wavelength sensitivity which varies as a sample ofinfrared energy is taken from the material sample S. Thus, thisembodiment permits the use of only one detector 28, the output signal ofwhich varies in wavelength specificity over time. The output signal canbe de-multiplexed based on the wavelength sensitivities induced by theFabry-Perot interferometer, to provide a multiple-wavelength profile ofthe infrared energy emitted by the material sample S. In thisembodiment, the optical mixer 20 may be omitted, as only one detector 28need be employed.

[0104] In still other embodiments, the array of filters 24 may comprisea filter wheel that rotates different filters with varying wavelengthsensitivities over a single detector 24. Alternatively, anelectronically tunable infrared filter may be employed in a mannersimilar to the Fabry-Perot interferometer discussed above, to providewavelength sensitivity which varies during the detection process. Ineither of these embodiments, the optical mixer 20 may be omitted, asonly one detector 28 need be employed.

[0105] f. Detectors

[0106] The detectors 28 may comprise any detector type suitable forsensing infrared energy, preferably in the mid-infrared wavelengths. Forexample, the detectors 28 may comprise mercury-cadmium-telluride (MCT)detectors. A detector such as a Fermionics (Simi Valley, Calif.) modelPV-9.1 with a PVA481-1 pre-amplifier is acceptable. Similar units fromother manufacturers such as Graseby (Tampa, Fla.) can be substituted.Other suitable components for use as the detectors 28 includepyroelectric detectors, thermopiles, bolometers, silicon microbolometersand lead-salt focal plane arrays.

[0107] g. Control System

[0108]FIG. 7 depicts the control system 30 in greater detail, as well asthe interconnections between the control system and other relevantportions of the noninvasive system. The control system includes atemperature control subsystem and a data acquisition subsystem.

[0109] In the temperature control subsystem, temperature sensors (suchas RTDs and/or thermistors) located in the window assembly 12 provide awindow temperature signal to a synchronous analog-to-digital conversionsystem 70 and an asynchronous analog-to-digital conversion system 72.The A/D systems 70, 72 in turn provide a digital window temperaturesignal to a digital signal processor (DSP) 74. The processor 74 executesa window temperature control algorithm and determines appropriatecontrol inputs for the heater layer 34 of the window assembly 12 and/orfor the cooling system 14, based on the information contained in thewindow temperature signal. The processor 74 outputs one or more digitalcontrol signals to a digital-to-analog conversion system 76 which inturn provides one or more analog control signals to current drivers 78.In response to the control signal(s), the current drivers 78 regulatethe power supplied to the heater layer 34 and/or to the cooling system14. In one embodiment, the processor 74 provides a control signalthrough a digital I/O device 77 to a pulse-width modulator (PWM) control80, which provides a signal that controls the operation of the currentdrivers 78. Alternatively, a low-pass filter (not shown) at the outputof the PWM provides for continuous operation of the current drivers 78.

[0110] In another embodiment, temperature sensors may be located at thecooling system 14 and appropriately connected to the A/D system(s) andprocessor to provide closed-loop control of the cooling system as well.

[0111] In yet another embodiment, a detector cooling system 82 islocated in thermally conductive relation to one or more of the detectors28. The detector cooling system 82 may comprise any of the devicesdisclosed above as comprising the cooling system 14, and preferablycomprises a Peltier-type thermoelectric device. The temperature controlsubsystem may also include temperature sensors, such as RTDs and/orthermistors, located in or adjacent to the detector cooling system 82,and electrical connections between these sensors and the asynchronousA/D system 72. The temperature sensors of the detector cooling system 82provide detector temperature signals to the processor 74. In oneembodiment, the detector cooling system 82 operates independently of thewindow temperature control system, and the detector cooling systemtemperature signals are sampled using the asynchronous A/D system 72. Inaccordance with the temperature control algorithm, the processor 74determines appropriate control inputs for the detector cooling system82, based on the information contained in the detector temperaturesignal. The processor 74 outputs digital control signals to the D/Asystem 76 which in turn provides analog control signals to the currentdrivers 78. In response to the control signals, the current drivers 78regulate the power supplied to the detector cooling system 14. In oneembodiment, the processor 74 also provides a control signal through thedigital I/O device 77 and the PWM control 80, to control the operationof the detector cooling system 82 by the current drivers 78.Alternatively, a low-pass filter (not shown) at the output of the PWMprovides for continuous operation of the current drivers 78.

[0112] In the data acquisition subsystem, the detectors 28 respond tothe infrared energy E incident thereon by passing one or more analogdetector signals to a preamp 84. The preamp 84 amplifies the detectorsignals and passes them to the synchronous A/D system 70, which convertsthe detector signals to digital form and passes them to the processor74. The processor 74 determines the concentrations of the analyte(s) ofinterest, based on the detector signals and a concentration-analysisalgorithm and/or phase/concentration regression model stored in a memorymodule 88. The concentration-analysis algorithm and/orphase/concentration regression model may be developed according to anyof the analysis methodologies discussed herein. The processor maycommunicate the concentration results and/or other information to adisplay controller 86, which operates a display (not shown), such as anLCD display, to present the information to the user.

[0113] A watchdog timer 94 may be employed to ensure that the processor74 is operating correctly. If the watchdog timer 94 does not receive asignal from the processor 74 within a specified time, the watchdog timer94 resets the processor 74. The control system may also include a JTAGinterface 96 to enable testing of the noninvasive system 10.

[0114] In one embodiment, the synchronous A/D system 70 comprises a20-bit, 14 channel system, and the asynchronous A/D system 72 comprisesa 16-bit, 16 channel system. The preamp may comprise a 12-channel preampcorresponding to an array of 12 detectors 28.

[0115] The control system may also include a serial port 90 or otherconventional data port to permit connection to a personal computer 92.The personal computer can be employed to update the algorithm(s) and/orphase/concentration regression model(s) stored in the memory module 88,or to download a compilation of analyte-concentration data from thenoninvasive system. A real-time clock or other timing device may beaccessible by the processor 74 to make any time-dependent calculationswhich may be desirable to a user.

[0116] 2. Analysis Methodology

[0117] The detector(s) 28 of the noninvasive system 10 are used todetect the infrared energy emitted by the material sample S in variousdesired wavelengths. At each measured wavelength, the material sample Semits infrared energy at an intensity which varies over time. Thetime-varying intensities arise largely in response to the use of thewindow assembly 12 (including its heater layer 34) and the coolingsystem 14 to induce a thermal gradient in the material sample S. As usedherein, “thermal gradient” is a broad term and is used in its ordinarysense and refers, without limitation, to a difference in temperatureand/or thermal energy between different locations, such as differentdepths, of a material sample, which can be induced by any suitablemethod of increasing or decreasing the temperature and/or thermal energyin one or more locations of the sample. As will be discussed in detailbelow, the concentration of an analyte of interest (such as glucose) inthe material sample S can be determined with a device such as thenoninvasive system 10, by comparing the time-varying intensity profilesof the various measured wavelengths.

[0118] Analysis methodologies are discussed herein within the context ofdetecting the concentration of glucose within a material sample, such asa tissue sample, which includes a large proportion of water. However, itwill evident that these methodologies are not limited to this contextand may be applied to the detection of a wide variety of analytes withina wide variety of sample types. It should also be understood that othersuitable analysis methodologies and suitable variations of the disclosedmethodologies may be employed in operating an analyte detection system,such as the noninvasive system 10.

[0119] As shown in FIG. 8, a first reference signal P may be measured ata first reference wavelength. The first reference signal P is measuredat a wavelength where water strongly absorbs (e.g., 2.9 μm or 6.1 μm).Because water strongly absorbs radiation at these wavelengths, thedetector signal intensity is reduced at those wavelengths. Moreover, atthese wavelengths water absorbs the photon emissions emanating from deepinside the sample. The net effect is that a signal emitted at thesewavelengths from deep inside the sample is not easily detected. Thefirst reference signal P is thus a good indicator of thermal-gradienteffects near the sample surface and may be known as a surface referencesignal. This signal may be calibrated and normalized, in the absence ofheating or cooling applied to the sample, to a baseline value of 1. Forgreater accuracy, more than one first reference wavelength may bemeasured. For example, both 2.9 μm and 6.1 μm may be chosen as firstreference wavelengths.

[0120] As further shown in FIG. 8, a second reference signal R may alsobe measured. The second signal R may be measured at a wavelength wherewater has very low absorbance (e.g., 3.6 μm or 4.2 μm). This secondreference signal R thus provides the analyst with information concerningthe deeper regions of the sample, whereas the first signal P providesinformation concerning the sample surface. This signal may also becalibrated and normalized, in the absence of heating or cooling appliedto the sample, to a baseline value of 1. As with the first (surface)reference signal P, greater accuracy may be obtained by using more thanone second (deep) reference signal R.

[0121] In order to determine analyte concentration, a third (analytical)signal Q is also measured. This signal is measured at an IR absorbancepeak of the selected analyte. The IR absorbance peaks for glucose are inthe range of about 6.5 μm to 11.0 μm. This detector signal may also becalibrated and normalized, in the absence of heating or cooling appliedto the material sample S, to a baseline value of 1. As with thereference signals P, R, the analytical signal Q may be measured at morethan one absorbance peak.

[0122] Optionally, or additionally, reference signals may be measured atwavelengths that bracket the analyte absorbance peak. These signals maybe advantageously monitored at reference wavelengths which do notoverlap the analyte absorbance peaks. Further, it is advantageous tomeasure reference wavelengths at absorbance peaks which do not overlapthe absorbance peaks of other possible constituents contained in thesample.

[0123] a. Basic Thermal Gradient

[0124] As further shown in FIG. 8, the signal intensities P, Q, R areshown initially at the normalized baseline signal intensity of 1. Thisof course reflects the baseline radiative behavior of a test sample inthe absence of applied heating or cooling. At a time t_(C), the surfaceof the sample is subjected to a temperature event which induces athermal gradient in the sample. The gradient can be induced by heatingor cooling the sample surface. The example shown in FIG. 8 uses cooling,for example, using a 10° C. cooling event. In response to the coolingevent, the intensities of the detector signals P, Q, R decrease overtime.

[0125] Since the cooling of the sample is neither uniform norinstantaneous, the surface cools before the deeper regions of the samplecool. As each of the signals P, Q, R drop in intensity, a patternemerges. Signal intensity declines as expected, but as the signals P, Q,R reach a given amplitude value (or series of amplitude values: 150,152, 154, 156, 158), certain temporal effects are noted. After thecooling event is induced at t_(C), the first (surface) reference signalP declines in amplitude most rapidly, reaching a checkpoint 150 first,at time t_(P). This is due to the fact that the first reference signal Pmirrors the sample's radiative characteristics near the surface of thesample. Since the sample surface cools before the underlying regions,the surface (first) reference signal P drops in intensity first.

[0126] Simultaneously, the second reference signal R is monitored. Sincethe second reference signal R corresponds to the radiationcharacteristics of deeper regions of the sample, which do not cool asrapidly as the surface (due to the time needed for the surface coolingto propagate into the deeper regions of the sample), the intensity ofsignal R does not decline until slightly later. Consequently, the signalR does not reach the magnitude 150 until some later time t_(R). In otherwords, there exists a time delay between the time t_(P) at which theamplitude of the first reference signal P reaches the checkpoint 150 andthe time t_(R) at which the second reference signal R reaches the samecheckpoint 150. This time delay can be expressed as a phase differenceΦ(λ). Additionally, a phase difference may be measured between theanalytical signal Q and either or both reference signals P, R.

[0127] As the concentration of analyte increases, the amount ofabsorbance at the analytical wavelength increases. This reduces theintensity of the analytical signal Q in a concentration-dependent way.Consequently, the analytical signal Q reaches intensity 150 at someintermediate time t_(Q). The higher the concentration of analyte, themore the analytical signal Q shifts to the left in FIG. 8. As a result,with increasing analyte concentration, the phase difference Φ(λ)decreases relative to the first (surface) reference signal P andincreases relative to the second (deep tissue) reference signal R. Thephase difference(s) Φ(λ) are directly related to analyte concentrationand can be used to make accurate determinations of analyteconcentration.

[0128] The phase difference Φ(λ) between the first (surface) referencesignal P and the analytical signal Q is represented by the equation:

Φ(λ)=|t _(P) −t _(Q)|

[0129] The magnitude of this phase difference decreases with increasinganalyte concentration.

[0130] The phase difference Φ(λ) between the second (deep tissue)reference signal R and the analytical signal Q signal is represented bythe equation:

Φ(λ)=|t _(Q) −t _(R)|

[0131] The magnitude of this phase difference increases with increasinganalyte concentration.

[0132] Accuracy may be enhanced by choosing several checkpoints, forexample, 150, 152, 154, 156, and 158 and averaging the phase differencesobserved at each checkpoint. The accuracy of this method may be furtherenhanced by integrating the phase difference(s) continuously over theentire test period. Because in this example only a single temperatureevent (here, a cooling event) has been induced, the sample reaches a newlower equilibrium temperature and the signals stabilize at a newconstant level I_(F). Of course, the method works equally well withthermal gradients induced by heating or by the application orintroduction of other forms of energy, such as but not limited to light,radiation, chemically induced heat, friction and vibration.

[0133] This methodology is not limited to the determination of phasedifference. At any given time (for example, at a time t_(X)) theamplitude of the analytical signal Q may be compared to the amplitude ofeither or both of the reference signals P, R. The difference inamplitude may be observed and processed to determine analyteconcentration.

[0134] This method, the variants disclosed herein, and the apparatusdisclosed as suitable for application of the method(s), are not limitedto the detection of in-vivo glucose concentration. The method anddisclosed variants and apparatus may be used on human, animal, or evenplant subjects, or on organic or inorganic compositions in a non-medicalsetting. The method may be used to take measurements of in-vivo orin-vitro samples of virtually any kind. The method is useful formeasuring the concentration of a wide range of additional chemicalanalytes, including but not limited to, glucose, ethanol, insulin,water, carbon dioxide, blood oxygen, cholesterol, bilirubin, ketones,fatty acids, lipoproteins, albumin, urea, creatinine, white blood cells,red blood cells, hemoglobin, oxygenated hemoglobin, carboxyhemoglobin,organic molecules, inorganic molecules, pharmaceuticals, cytochrome,various proteins and chromophores, microcalcifications, hormones, aswell as other chemical compounds. To detect a given analyte, one needsonly to select appropriate analytical and reference wavelengths.

[0135] The method is adaptable and may be used to determine chemicalconcentrations in samples of body fluids (e.g., blood, urine or saliva)once they have been extracted from a patient. In fact, the method may beused for the measurement of in-vitro samples of virtually any kind.

[0136] b. Modulated Thermal Gradient

[0137] In some embodiments of the methodology described above, aperiodically modulated thermal gradient can be employed to make accuratedeterminations of analyte concentration.

[0138] As previously shown in FIG. 8, once a thermal gradient is inducedin the sample, the reference and analytical signals P, Q, R fall out ofphase with respect to each other. This phase difference Φ(λ) is presentwhether the thermal gradient is induced through heating or cooling. Byalternatively subjecting the test sample to cyclic pattern of heating,cooling, or alternately heating and cooling, an oscillating thermalgradient may be induced in a sample for an extended period of time.

[0139] An oscillating thermal gradient is illustrated using asinusoidally modulated gradient. FIG. 9 depicts detector signalsemanating from a test sample. As with the methodology shown in FIG. 8,one or more reference signals J, L are measured. One or more analyticalsignals K are also monitored. These signals may be calibrated andnormalized, in the absence of heating or cooling applied to the sample,to a baseline value of 1. FIG. 9 shows the signals after normalization.At some time t_(C), a temperature event (e.g., cooling) is induced atthe sample surface. This causes a decline in the detector signal. Asshown in FIG. 8, the signals (P, Q, R) decline until the thermalgradient disappears and a new equilibrium detector signal I_(F) isreached. In the method shown in FIG. 9, as the gradient begins todisappear at a signal intensity 160, a heating event, at a time t_(W),is induced in the sample surface. As a result the detector outputsignals J, K, L will rise as the sample temperature rises. At some latertime t_(C2), another cooling event is induced, causing the temperatureand detector signals to decline. This cycle of cooling and heating maybe repeated over a time interval of arbitrary length. Moreover, if thecooling and heating events are timed properly, a periodically modulatedthermal gradient may be induced in the test sample.

[0140] As previously explained in the discussions relating to FIG. 8,the phase difference Φ(λ) may be measured and used to determine analyteconcentration. FIG. 9 shows that the first (surface) reference signal Jdeclines and rises in intensity first. The second (deep tissue)reference signal L declines and rises in a time-delayed manner relativeto the first reference signal J. The analytical signal K exhibits atime/phase delay dependent on the analyte concentration. With increasingconcentration, the analytical signal K shifts to the left in FIG. 9. Aswith FIG. 8, the phase difference Φ(λ) may be measured. For example, aphase difference Φ(λ) between the second reference signal L and theanalytical signal K, may be measured at a set amplitude 162 as shown inFIG. 9. Again, the magnitude of the phase signal reflects the analyteconcentration of the sample.

[0141] The phase-difference information compiled by any of themethodologies disclosed herein can correlated by the control system 30(see FIG. 1) with previously determined phase-difference information todetermine the analyte concentration in the sample. This correlationcould involve comparison of the phase-difference information receivedfrom analysis of the sample, with a data set containing thephase-difference profiles observed from analysis of wide variety ofstandards of known analyte concentration. In one embodiment, aphase/concentration curve or regression model is established by applyingregression techniques to a set of phase-difference data observed instandards of known analyte concentration. This curve is used to estimatethe analyte concentration in a sample based on the phase-differenceinformation received from the sample.

[0142] Advantageously, the phase difference Φ(λ) may be measuredcontinuously throughout the test period. The phase-differencemeasurements may be integrated over the entire test period for anextremely accurate measure of phase difference Φ(λ). Accuracy may alsobe improved by using more than one reference signal and/or more than oneanalytical signal.

[0143] As an alternative or as a supplement to measuring phasedifference(s), differences in amplitude between the analytical andreference signal(s) may be measured and employed to determine analyteconcentration. Additional details relating to this technique and notnecessary to repeat here may be found in the Assignee's U.S. patentapplication Ser. No. 09/538,164, incorporated by reference below.

[0144] Additionally, these methods may be advantageously employed tosimultaneously measure the concentration of one or more analytes. Bychoosing reference and analyte wavelengths that do not overlap, phasedifferences can be simultaneously measured and processed to determineanalyte concentrations. Although FIG. 9 illustrates the method used inconjunction with a sinusoidally modulated thermal gradient, theprinciple applies to thermal gradients conforming to any periodicfunction. In more complex cases, analysis using signal processing withFourier transforms or other techniques allows accurate determinations ofphase difference Φ(λ) and analyte concentration.

[0145] As shown in FIG. 10, the magnitude of the phase differences maybe determined by measuring the time intervals between the amplitudepeaks (or troughs) of the reference signals J, L and the analyticalsignal K. Alternatively, the time intervals between the “zero crossings”(the point at which the signal amplitude changes from positive tonegative, or negative to positive) may be used to determine the phasedifference between the analytical signal K and the reference signals J,L. This information is subsequently processed and a determination ofanalyte concentration may then be made. This particular method has theadvantage of not requiring normalized signals.

[0146] As a further alternative, two or more driving frequencies may beemployed to determine analyte concentrations at selected depths withinthe sample. A slow (e.g., 1 Hz) driving frequency creates a thermalgradient which penetrates deeper into the sample than the gradientcreated by a fast (e.g., 3 Hz) driving frequency. This is because theindividual heating and/or cooling events are longer in duration wherethe driving frequency is lower. Thus, the use of a slow drivingfrequency provides analyte-concentration information from a deeper“slice” of the sample than does the use of a fast driving frequency.

[0147] It has been found that when analyzing a sample of human skin, atemperature event of 10° C. creates a thermal gradient which penetratesto a depth of about 150 μm, after about 500 ms of exposure.Consequently, a cooling/heating cycle or driving frequency of 1 Hzprovides information to a depth of about 150 μm. It has also beendetermined that exposure to a temperature event of 10° C. for about 167ms creates a thermal gradient that penetrates to a depth of about 50 μm.Therefore, a cooling/heating cycle of 3 Hz provides information to adepth of about 50 μm. By subtracting the detector signal informationmeasured at a 3 Hz driving frequency from the detector signalinformation measured at a 1 Hz driving frequency, one can determine theanalyte concentration(s) in the region of skin between 50 and 150 μm. Ofcourse, a similar approach can be used to determine analyteconcentrations at any desired depth range within any suitable type ofsample.

[0148] As shown in FIG. 11, alternating deep and shallow thermalgradients may be induced by alternating slow and fast drivingfrequencies. As with the methods described above, this variation alsoinvolves the detection and measurement of phase differences Φ(λ) betweenreference signals G, G′ and analytical signals H, H′. Phase differencesare measured at both fast (e.g., 3 Hz) and slow (e.g., 1 Hz) drivingfrequencies. The slow driving frequency may continue for an arbitrarilychosen number of cycles (in region SL₁), for example, two full cycles.Then the fast driving frequency is employed for a selected duration, inregion F₁. The phase difference data is compiled in the same manner asdisclosed above. In addition, the fast frequency (shallow sample) phasedifference data may be subtracted from the slow frequency (deep sample)data to provide an accurate determination of analyte concentration inthe region of the sample between the gradient penetration depthassociated with the fast driving frequency and that associated with theslow driving frequency.

[0149] The driving frequencies (e.g., 1 Hz and 3 Hz) can be multiplexedas shown in FIG. 12. The fast (3 Hz) and slow (1 Hz) driving frequenciescan be superimposed rather than sequentially implemented. Duringanalysis, the data can be separated by frequency (using Fouriertransform or other techniques) and independent measurements of phasedelay at each of the driving frequencies may be calculated. Onceresolved, the two sets of phase delay data are processed to determineabsorbance and analyte concentration.

[0150] Additional details not necessary to repeat here may be found inU.S. Pat. No. 6,198,949, titled SOLID-STATE NON-INVASIVE INFRAREDABSORPTION SPECTROMETER FOR THE GENERATION AND CAPTURE OF THERMALGRADIENT SPECTRA FROM LIVING TISSUE, issued Mar. 6, 2001; U.S. Pat. No.6,161,028, titled METHOD FOR DETERMINING ANALYTE CONCENTRATION USINGPERIODIC TEMPERATURE MODULATION AND PHASE DETECTION, issued Dec. 12,2000; U.S. Pat. No. 5,877,500, titled MULTICHANNEL INFRARED DETECTORWITH OPTICAL CONCENTRATORS FOR EACH CHANNEL, issued on Mar. 2, 1999;U.S. patent application Ser. No. 09/538,164, filed Mar. 30, 2000 andtitled METHOD AND APPARATUS FOR DETERMINING ANALYTE CONCENTRATION USINGPHASE AND MAGNITUDE DETECTION OF A RADIATION TRANSFER FUNCTION; U.S.Provisional Patent Application No. 60/336,404, filed Oct. 29, 2001,titled WINDOW ASSEMBLY; U.S. Provisional Patent Application No.60/340,435, filed Dec. 12, 2001, titled CONTROL SYSTEM FOR BLOODCONSTITUENT MONITOR; U.S. Provisional Patent Application No. 60/340,654,filed Dec. 12, 2001, titled SYSTEM AND METHOD FOR CONDUCTING ANDDETECTING INFRARED RADIATION; U.S. Provisional Patent Application No.60/336,294, filed Oct. 29, 2001, titled METHOD AND DEVICE FOR INCREASINGACCURACY OF BLOOD CONSTITUENT MEASUREMENT; and U.S. Provisional PatentApplication No. 60/339,116, filed Nov. 7, 2001, titled METHOD ANDAPPARATUS FOR IMPROVING CLINICALLY SIGNIFICANT ACCURACY OF ANALYTEMEASUREMENTS. The entire disclosure of all of the above-mentionedpatents, patent applications and publications is hereby incorporated byreference herein and made a part of this specification.

[0151] B. Whole-Blood Detection System

[0152]FIG. 13 is a schematic view of a reagentless whole-blood analytedetection system 200 (hereinafter “whole-blood system”) in a preferredconfiguration. The whole-blood system 200 may comprise a radiationsource 220, a filter 230, a cuvette 240 that includes a sample cell 242,and a radiation detector 250. The whole-blood system 200 preferably alsocomprises a signal processor 260 and a display 270. Although a cuvette240 is shown here, other sample elements, as described below, could alsobe used in the system 200. The whole-blood system 200 can also comprisea sample extractor 280, which can be used to access bodily fluid from anappendage, such as the finger 290, forearm, or any other suitablelocation.

[0153] As used herein, the terms “whole-blood analyte detection system”and “whole-blood system” are broad, synonymous terms and are used intheir ordinary sense and refer, without limitation, to analyte detectiondevices which can determine the concentration of an analyte in amaterial sample by passing electromagnetic radiation into the sample anddetecting the absorbance of the radiation by the sample. As used herein,the term “wholeblood” is a broad term and is used in its ordinary senseand refers, without limitation, to blood that has been withdrawn from apatient but that has not been otherwise processed, e.g., it has not beenhemolysed, lyophilized, centrifuged, or separated in any other manner,after being removed from the patient. Whole-blood may contain amounts ofother fluids, such as interstitial fluid or intracellular fluid, whichmay enter the sample during the withdrawal process or are naturallypresent in the blood. It should be understood, however, that thewhole-blood system 200 disclosed herein is not limited to analysis ofwhole-blood, as the whole-blood system 10 may be employed to analyzeother substances, such as saliva, urine, sweat, interstitial fluid,intracellular fluid, hemolysed, lyophilized, or centrifuged blood or anyother organic or inorganic materials.

[0154] The whole-blood system 200 may comprise a near-patient testingsystem. As used herein, “near-patient testing system” is a broad termand is used in its ordinary sense, and includes, without limitation,test systems that are configured to be used where the patient is ratherthan exclusively in a laboratory, e.g., systems that can be used at apatient's home, in a clinic, in a hospital, or even in a mobileenvironment. Users of near-patient testing systems can include patients,family members of patients, clinicians, nurses, or doctors. A“near-patient testing system” could also include a “point-of-care”system.

[0155] The whole-blood system 200 may in one embodiment be configured tobe operated easily by the patient or user. As such, the system 200 ispreferably a portable device. As used herein, “portable” is a broad termand is used in its ordinary sense and means, without limitation, thatthe system 200 can be easily transported by the patient and used whereconvenient. For example, the system 200 is advantageously small. In onepreferred embodiment, the system 200 is small enough to fit into a purseor backpack. In another embodiment, the system 200 is small enough tofit into a pants pocket. In still another embodiment, the system 200 issmall enough to be held in the palm of a hand of the user.

[0156] Some of the embodiments described herein employ a sample elementto hold a material sample, such as a sample of biological fluid. As usedherein, “sample element” is a broad term and is used in its ordinarysense and includes, without limitation, structures that have a samplecell and at least one sample cell wall, but more generally includes anyof a number of structures that can hold, support or contain a materialsample and that allow electromagnetic radiation to pass through a sampleheld, supported or contained thereby; e.g., a cuvette, test strip, etc.As used herein, the term “disposable” when applied to a component, suchas a sample element, is a broad term and is used in its ordinary senseand means, without limitation, that the component in question is used afinite number of times and then discarded. Some disposable componentsare used only once and then discarded. Other disposable components areused more than once and then discarded.

[0157] The radiation source 220 of the whole-blood system 200 emitselectromagnetic radiation in any of a number of spectral ranges, e.g.,within infrared wavelengths; in the mid-infrared wavelengths; aboveabout 0.8 μm; between about 5.0 μm and about 20.0 m; and/or betweenabout 5.25 μm and about 12.0 μm. However, in other embodiments thewhole-blood system 200 may employ a radiation source 220 which emits inwavelengths found anywhere from the visible spectrum through themicrowave spectrum, for example anywhere from about 0.4 μm to greaterthan about 100 μm. In still further embodiments the radiation sourceemits electromagnetic radiation in wavelengths between about 3.5 μm andabout 14 μm, or between about 0.8 μm and about 2.5 μm, or between about2.5 μm and about 20 μm, or between about 20 μm and about 100 μm, orbetween about 6.85 μm and about 10.10 μm.

[0158] The radiation emitted from the source 220 is in one embodimentmodulated at a frequency between about one-half hertz and about onehundred hertz, in another embodiment between about 2.5 hertz and about7.5 hertz, in still another embodiment at about 50 hertz, and in yetanother embodiment at about 5 hertz. With a modulated radiation source,ambient light sources, such as a flickering fluorescent lamp, can bemore easily identified and rejected when analyzing the radiationincident on the detector 250. One source that is suitable for thisapplication is produced by ION OPTICS, INC. and sold under the partnumber NL5LNC.

[0159] The filter 230 permits electromagnetic radiation of selectedwavelengths to pass through and impinge upon the cuvette/sample element240. Preferably, the filter 230 permits radiation at least at about thefollowing wavelengths to pass through to the cuvette/sample element:3.9, 4.0 μm, 4.05 μm, 4.2 μm, 4.75, 4.95 μm, 5.25 μm, 6.12 μm, 7.4 μm,8.0 μm, 8.45 μm, 9.25 μm, 9.5 μm, 9.65 μm, 10.4 μm, 12.2 μm. In anotherembodiment, the filter 230 permits radiation at least at about thefollowing wavelengths to pass through to the cuvette/sample element:5.25 μm, 6.12 μm, 6.8 μm, 8.03 μm, 8.45 μm, 9.25 μm, 9.65 μm, 10.4 μm,12 μm. In still another embodiment, the filter 230 permits radiation atleast at about the following wavelengths to pass through to thecuvette/sample element: 6.85 μm, 6.97 μm, 7.39 μm, 8.23 μm, 8.62 μm,9.02 μm, 9.22 μm, 9.43 μm, 9.62 μm, and 10.10 μm. The sets ofwavelengths recited above correspond to specific embodiments within thescope of this disclosure. Furthermore, other subsets of the foregoingsets or other combinations of wavelengths can be selected. Finally,other sets of wavelengths can be selected within the scope of thisdisclosure based on cost of production, development time, availability,and other factors relating to cost, manufacturability, and time tomarket of the filters used to generate the selected wavelengths, and/orto reduce the total number of filters needed.

[0160] In one embodiment, the filter 230 is capable of cycling itspassband among a variety of narrow spectral bands or a variety ofselected wavelengths. The filter 230 may thus comprise a solid-statetunable infrared filter, such as that available from ION OPTICS INC. Thefilter 230 could also be implemented as a filter wheel with a pluralityof fixed-passband filters mounted on the wheel, generally perpendicularto the direction of the radiation emitted by the source 220. Rotation ofthe filter wheel alternately presents filters that pass radiation atwavelengths that vary in accordance with the filters as they passthrough the field of view of the detector 250.

[0161] The detector 250 preferably comprises a 3 mm long by 3 mm widepyroelectric detector. Suitable examples are produced by DIAS AngewandteSensorik GmbH of Dresden, Germany, or by BAE Systems (such as its TGSmodel detector). The detector 250 could alternatively comprise athermopile, a bolometer, a silicon microbolometer, a lead-salt focalplane array, or a mercury-cadmium-telluride (MCT) detector. Whicheverstructure is used as the detector 250, it is desirably configured torespond to the radiation incident upon its active surface 254 to produceelectrical signals that correspond to the incident radiation.

[0162] In one embodiment, the sample element comprises a cuvette 240which in turn comprises a sample cell 242 configured to hold a sample oftissue and/or fluid (such as whole-blood, blood components, interstitialfluid, intercellular fluid, saliva, urine, sweat and/or other organic orinorganic materials) from a patient within its sample cell. The cuvette240 is installed in the whole-blood system 200 with the sample cell 242located at least partially in the optical path 243 between the radiationsource 220 and the detector 250. Thus, when radiation is emitted fromthe source 220 through the filter 230 and the sample cell 242 of thecuvette 240, the detector 250 detects the radiation signal strength atthe wavelength(s) of interest. Based on this signal strength, the signalprocessor 260 determines the degree to which the sample in the cell 242absorbs radiation at the detected wavelength(s). The concentration ofthe analyte of interest is then determined from the absorption data viaany suitable spectroscopic technique.

[0163] As shown in FIG. 13, the whole-blood system 200 can also comprisea sample extractor 280. As used herein, the term “sample extractor” is abroad term and is used in its ordinary sense and refers, withoutlimitation, to any device which is suitable for drawing a samplematerial, such as whole-blood, other bodily fluids, or any other samplematerial, through the skin of a patient. In various embodiments, thesample extractor may comprise a lance, laser lance, iontophoreticsampler, gas-jet, fluid-jet or particle-jet perforator, ultrasonicenhancer (used with or without a chemical enhancer), or any othersuitable device.

[0164] As shown in FIG. 13, the sample extractor 280 could form anopening in an appendage, such as the finger 290, to make whole-bloodavailable to the cuvette 240. It should be understood that otherappendages could be used to draw the sample, including but not limitedto the forearm. With some embodiments of the sample extractor 280, theuser forms a tiny hole or slice through the skin, through which flows asample of bodily fluid such as whole-blood. Where the sample extractor280 comprises a lance (see FIG. 14), the sample extractor 280 maycomprise a sharp cutting implement made of metal or other rigidmaterials. One suitable laser lance is the Lasette Plus® produced byCell Robotics International, Inc. of Albuquerque, N. Mex. If a laserlance, iontophoretic sampler, gas-jet or fluid-jet perforator is used asthe sample extractor 280, it could be incorporated into the whole-bloodsystem 200 (see FIG. 13), or it could be a separate device.

[0165] Additional information on laser lances can be found in U.S. Pat.No. 5,908,416, issued Jun. 1, 1999, titled LASER DERMAL PERFORATOR; theentirety of this patent is hereby incorporated by reference herein andmade a part of this specification. One suitable gas-jet, fluid-jet orparticle-jet perforator is disclosed in U.S. Pat. No. 6,207,400, issuedMar. 27, 2001, titled NON- OR MINIMALLY INVASIVE MONITORING METHODSUSING PARTICLE DELIVERY METHODS; the entirety of this patent is herebyincorporated by reference herein and made a part of this specification.One suitable iontophoretic sampler is disclosed in U.S. Pat. No.6,298,254, issued Oct. 2, 2001, titled DEVICE FOR SAMPLING SUBSTANCESUSING ALTERNATING POLARITY OF IONTOPHORETIC CURRENT; the entirety ofthis patent is hereby incorporated by reference herein and made a partof this specification. One suitable ultrasonic enhancer, and chemicalenhancers suitable for use therewith, are disclosed in U.S. Pat. No.5,458,140, titled ENHANCEMENT OF TRANSDERMAL MONITORING APPLICATIONSWITH ULTRASOUND AND CHEMICAL ENHANCERS, issued Oct. 17, 1995, the entiredisclosure of which is hereby incorporated by reference and made a partof this specification.

[0166]FIG. 14 shows one embodiment of a sample element, in the form of acuvette 240, in greater detail. The cuvette 240 further comprises asample supply passage 248, a pierceable portion 249, a first window 244,and a second window 246, with the sample cell 242 extending between thewindows 244, 246. In one embodiment, the cuvette 240 does not have asecond window 246. The first window 244 (or second window 246) is oneform of a sample cell wall; in other embodiments of the sample elementsand cuvettes disclosed herein, any sample cell wall may be used that atleast partially contains, holds or supports a material sample, such as abiological fluid sample, and which is transmissive of at least somebands of electromagnetic radiation, and which may but need not betransmissive of electromagnetic radiation in the visible range. Thepierceable portion 249 is an area of the sample supply passage 248 thatcan be pierced by suitable embodiments of the sample extractor 280.Suitable embodiments of the sample extractor 280 can pierce the portion249 and the appendage 290 to create a wound in the appendage 290 and toprovide an inlet for the blood or other fluid from the wound to enterthe cuvette 240. (The sample extractor 280 is shown on the opposite sideof the sample element in FIG. 14, as compared to FIG. 13, as it maypierce the portion 249 from either side.)

[0167] The windows 244, 246 are preferably optically transmissive in therange of electromagnetic radiation that is emitted by the source 220, orthat is permitted to pass through the filter 230. In one embodiment, thematerial that makes up the windows 244, 246 is completely transmissive,i.e., it does not absorb any of the electromagnetic radiation from thesource 220 and filter 230 that is incident upon it. In anotherembodiment, the material of the windows 244, 246 has some absorption inthe electromagnetic range of interest, but its absorption is negligible.In yet another embodiment, the absorption of the material of the windows244, 246 is not negligible, but it is known and stable for a relativelylong period of time. In another embodiment, the absorption of thewindows 244, 246 is stable for only a relatively short period of time,but the whole-blood system 200 is configured to observe the absorptionof the material and eliminate it from the analyte measurement before thematerial properties can change measurably.

[0168] The windows 244, 246 are made of polypropylene in one embodiment.In another embodiment, the windows 244, 246 are made of polyethylene.Polyethylene and polypropylene are materials having particularlyadvantageous properties for handling and manufacturing, as is known inthe art. Also, polypropylene can be arranged in a number of structures,e.g., isotactic, atactic and syndiotactic, which may enhance the flowcharacteristics of the sample in the sample element. Preferably thewindows 244, 246 are made of durable and easily manufactureablematerials, such as the above-mentioned polypropylene or polyethylene, orsilicon or any other suitable material. The windows 244, 246 can be madeof any suitable polymer, which can be isotactic, atactic or syndiotacticin structure.

[0169] The distance between the windows 244, 246 comprises an opticalpathlength and can be between about 1 μm and about 100 μm. In oneembodiment, the optical pathlength is between about 10 μm and about 40μm, or between about 25 μm and about 60 μm, or between about 30 μm andabout 50 μm. In still another embodiment, the optical pathlength isabout 25 μm. The transverse size of each of the windows 244, 246 ispreferably about equal to the size of the detector 250. In oneembodiment, the windows are round with a diameter of about 3 mm. In thisembodiment, where the optical pathlength is about 25 μm the volume ofthe sample cell 242 is about 0.177 μL. In one embodiment, the length ofthe sample supply passage 248 is about 6 mm, the height of the samplesupply passage 248 is about 1 mm, and the thickness of the sample supplypassage 248 is about equal to the thickness of the sample cell, e.g., 25μm. The volume of the sample supply passage is about 0.150 μL. Thus, thetotal volume of the cuvette 240 in one embodiment is about 0.327 μL. Ofcourse, the volume of the cuvette 240/sample cell 242/etc. can vary,depending on many variables, such as the size and sensitivity of thedetectors 250, the intensity of the radiation emitted by the source 220,the expected flow properties of the sample, and whether flow enhancers(discussed below) are incorporated into the cuvette 240. The transportof fluid to the sample cell 242 is achieved preferably through capillaryaction, but may also be achieved through wicking, or a combination ofwicking and capillary action.

[0170] FIGS. 15-17 depict another embodiment of a cuvette 305 that couldbe used in connection with the whole-blood system 200. The cuvette 305comprises a sample cell 310, a sample supply passage 315, an air ventpassage 320, and a vent 325. As best seen in FIGS. 16, 16A and 17, thecuvette also comprises a first sample cell window 330 having an innerside 332, and a second sample cell window 335 having an inner side 337.As discussed above, the window(s) 330/335 in some embodiments alsocomprise sample cell wall(s). The cuvette 305 also comprises an opening317 at the end of the sample supply passage 315 opposite the sample cell310. The cuvette 305 is preferably about ¼-⅛ inch wide and about ¾ inchlong; however, other dimensions are possible while still achieving theadvantages of the cuvette 305.

[0171] The sample cell 310 is defined between the inner side 332 of thefirst sample cell window 330 and the inner side 337 of the second samplecell window 335. The perpendicular distance T between the two innersides 332, 337 comprises an optical pathlength that can be between about1 μm and about 1.22 mm. The optical pathlength can alternatively bebetween about 1 μm and about 100 μm. The optical pathlength could stillalternatively be about 80 μm, but is preferably between about 10 μm andabout 50 μm. In another embodiment, the optical pathlength is about 25μm. The windows 330, 335 are preferably formed from any of the materialsdiscussed above as possessing sufficient radiation transmissivity. Thethickness of each window is preferably as small as possible withoutoverly weakening the sample cell 310 or cuvette 305.

[0172] Once a wound is made in the appendage 290, the opening 317 of thesample supply passage 315 of the cuvette 305 is placed in contact withthe fluid that flows from the wound. In another embodiment, the sampleis obtained without creating a wound, e.g. as is done with a salivasample. In that case, the opening 317 of the sample supply passage 315of the cuvette 305 is placed in contact with the fluid obtained withoutcreating a wound. The fluid is then transported through the samplesupply passage 315 and into the sample cell 310 via capillary action.The air vent passage 320 improves the capillary action by preventing thebuildup of air pressure within the cuvette and allowing the blood todisplace the air as the blood flows therein.

[0173] Other mechanisms may be employed to transport the sample to thesample cell 310. For example, wicking could be used by providing awicking material in at least a portion of the sample supply passage 315.In another variation, wicking and capillary action could be usedtogether to transport the sample to the sample cell 310. Membranes couldalso be positioned within the sample supply passage 315 to move theblood while at the same time filtering out components that mightcomplicate the optical measurement performed by the whole-blood system200.

[0174]FIGS. 16 and 16A depict one approach to constructing the cuvette305. In this approach, the cuvette 305 comprises a first layer 350, asecond layer 355, and a third layer 360. The second layer 355 ispositioned between the first layer 350 and the third layer 360. Thefirst layer 350 forms the first sample cell window 330 and the vent 325.As mentioned above, the vent 325 provides an escape for the air that isin the sample cell 310. While the vent 325 is shown on the first layer350, it could also be positioned on the third layer 360, or could be acutout in the second layer, and would then be located between the firstlayer 360 and the third layer 360 The third layer 360 forms the secondsample cell window 335.

[0175] The second layer 355 may be formed entirely of an adhesive thatjoins the first and third layers 350, 360. In other embodiments, thesecond layer may be formed from similar materials as the first and thirdlayers, or any other suitable material. The second layer 355 may also beformed as a carrier with an adhesive deposited on both sides thereof.The second layer 355 forms the sample supply passage 315, the air ventpassage 320, and the sample cell 310. The thickness of the second layer355 can be between about 1 μm and about 1.22 mm. This thickness canalternatively be between about 1 μm and about 100 μm. This thicknesscould alternatively be about 80 μm, but is preferably between about 10μm and about 50 μm. In another embodiment, the second layer thickness isabout 25 μm.

[0176] In other embodiments, the second layer 355 can be constructed asan adhesive film having a cutout portion to define the passages 315,320, or as a cutout surrounded by adhesive.

[0177] Further information can be found in U.S. patent application Ser.No. 10/055,875, filed Jan. 21, 2002, titled REAGENT-LESS WHOLE-BLOODGLUCOSE METER. The entire contents of this patent application isincorporated by reference herein and made a part of this specification.

II. Pathlength Insensitive Measurements

[0178] A variable optical pathlength algorithm facilitates pathlengthinsensitive measurements of blood constituents. This is to say that ifthe measurements are taken at the surface of a sample or at a depth of90% of the sample, inside a cuvette or in living tissue, the readingsare identical based on a mathematical correction process that appliesknown factors to the data before it is analyzed and a reading is givenfor the analyte in question.

[0179] Disclosed is a method that in certain embodiments accomplishesand solves all of the variable factors associated with taking bloodanalyte measurements in a blood sample, at any depth or thickness ofsample, and compensating for said variables.

[0180]FIG. 18 is a flow diagram of one embodiment of a method 500 forusing spectroscopy to determine an analyte concentration in a sample. Inan operational block 510, the method 500 comprises producing anabsorbance spectrum of the sample. In an operational block 520, themethod 500 further comprises shifting the absorbance spectrum to zero ina wavelength region. In an operational block 530, the method 500 furthercomprises subtracing a water or other substance contribution from theabsorbance spectrum.

[0181] A. Experimental Set-Up

[0182] In an exemplary embodiment, blood was simulated in its majorcomponents using bovine serum albumin (BSA) for total blood protein andsaline for serum. Infrared absorbance or optical density (OD) spectra ofthe samples were measured with a Fourier-Transform Infrared (FTIR)instrument from Perkin-Elmer, Inc. of Wellseley, Mass. As used herein,the term “optical density” or “OD” is synonymous with the term“absorbance.” Cuvette pathlength was set with different spacers betweenBaF₂ windows at 32 and 20 micrometers. The fringe pattern of the emptycuvette was used for calculation of the actual optical pathlength insidethe cuvette. Flexible tubing and the flow-through type of the cuvetteallowed for repeated filling with different solutions without changesbeing made to the experimental setup. Instrument drift and baselinedeviations were accounted for with saline reference measurements beforeand after sample measurements. A total of 100 scans were collected persample over a period of about 5 minutes. Scanned data were stored inASCII format and transferred to an electronic spreadsheet program (e.g.,Lotus 1-2-3 from IBM Corp. of Armonk, N.Y.) for evaluation.

[0183] B. Results

[0184] It was discovered that changes of the protein to water ratio,equivalent to changes of hematocrit in blood, produce at least 2isosbestic points on the wavelength scale, where the components proteinand water exhibit identical IR absorbance, one of them at about 4 μm andthe other at about 9.4 μm for BSA in water. As seen in FIG. 19 and FIG.20, the effective isosbestic point can be expected to be somewhatdifferent for different proteins in different solutions. The importantand unexpected aspect of this observation is that these wavelengthranges can be used to obtain a current measure of effective opticalpathlength in the filled cuvette, either before or during measurementsat other wavelength ranges. Such information is very useful insubsequent calculations for compensation of instrument-relatedpathlength non-linearities.

[0185] It was also discovered that after setting transmittance to zeroat one or both wavelengths ranges of high water absorbance at 6.08and/or 12.25 μm, thereby shifting absorbance to zero preferably at thelower isosbestic point, will result in spectral data at baseline. InFIG. 21, we can see the higher isosbestic point which may be used alsobut it is partially contaminated with absorbances of blood componentsthat are present at low concentration levels.

[0186] It was also discovered that the absorbance peak centered around4.7 μm, is almost entirely due to free water absorbance and can beadvantageously used for determination of free water in the sample: thecorrect subtraction of absorbance of a stored reference water across theentire wavelength range is achieved when there is zero residualabsorbance left between approximately 4.5 and 5 μm, shown in FIG. 21.

[0187] It was also discovered that prior knowledge of opticalpathlength, based on the total sample absorbance at the isosbestic pointas well as on water absorbance between approximately 4.5 and 5 μm,allows for the use of the correct reference water spectrum that iscompensated for non-linearities at all wavelengths. This is advantageousfor distortion-free presentation of final results.

[0188] It was also discovered that the absorbance peak centered around7.1 μm, is almost entirely due to free protein absorbance and can beadvantageously used for determination of free protein in the sample: thecorrect subtraction of absorbance of a stored reference hydrated proteinacross the entire wavelength range is achieved when there is zeroresidual absorbance left between approximately 7.0 and 7.2 μm oralternatively, at a different protein absorbance such as the range fromapproximately 7.9 to 8.1 μm or alternatively, a predefined residual at acombination of wavelength ranges, as shown in FIG. 22.

[0189] It was also discovered that prior knowledge of optical pathlengthbased on total sample absorbance at the isosbestic point as well as ontotal protein absorbance between approximately 7.0 and 7.2 μm, oralternatively, on a different protein absorbance such as the range fromapproximately 7.9 to 8.1 μm, allows for the use of the correct referenceprotein spectrum that is compensated for non-linearities at allwavelengths. This is advantageous for distortion-free presentation offinal results.

[0190]FIG. 23 shows that it was also discovered that repetition of thelast two steps of water and protein removal from the complete sample,will result in further removal of smaller residual interactivecomponents, most likely representing components of the boundary layerbetween water and protein.

[0191] It was also discovered that the resulting instrument driftcorrected, water-, protein-, interactive components-removed, and opticalpathlength and major component induced distortion-free spectral data canbe fitted with reference glucose spectral data at one or more glucoseabsorbance maxima such as 9.25 and 9.65 μm to yield a measure forglucose in the original sample. The residual absorbance after glucosespectral data removal may be used further for individual determinationof residual components. In certain embodiments, the residual componentsinclude high molecular weight substances, including but not limited to,other proteins, albumin, hemoglobin, fibrinogen, lipoproteins, andtrasferrin. In certain other embodiments, the residual componentsinclude low molecular weight substances, including but not limited to,urea, lactate, and vitamin C. The final glucose measure may be correctedfor the presence of such lower level potentially interfering substancesby subtracting reference spectra of specific substances, such as urea,from the residual data, as shown in FIG. 24.

[0192] It was also discovered that by following the definition ofglucose in blood, i.e. a measure of glucose per volume of sample, auseful measure for glucose concentration is obtained fromalgorithmically derived IR quantities as described above, by dividingthe final glucose quantity by total water, total protein oralternatively a combination of both.

[0193] The methods disclosed herein may be used in connection withapparatus and or methods disclosed in U.S. Pat. No. 6,198,949, titledSOLID-STATE NONINVASIVE INFRARED ABSORPTION SPECTROMETER FOR THEGENERATION AND CAPTURE OF THERMAL GRADIENT SPECTRA FROM LIVING TISSUE,issued Mar. 6, 2001; U.S. Pat. No. 6,161,028, titled METHOD FORDETERMINING ANALYTE CONCENTRATION USING PERIODIC TEMPERATURE MODULATIONAND PHASE DETECTION, issued Dec. 12, 2000; U.S. patent application Ser.No. 09/538,164, filed Mar. 30, 2000 and titled METHOD AND APPARATUS FORDETERMINING ANALYTE CONCENTRATION USING PHASE AND MAGNITUDE DETECTION OFA RADIATION TRANSFER FUNCTION; and WIPO PCT Publication No. WO 01/30236(corresponding to U.S. patent application Ser. No. 09/427,178),published May 3, 2001, titled SOLID-STATE NON-INVASIVE THERMAL CYCLINGSPECTROMETER. The entire disclosures of all of the above-mentionedpatents, patent applications and publications re hereby incorporated byreference herein and made a part of this specification.

[0194]FIG. 25 is a flow diagram of one embodiment of a method 600 ofproviding pathlength insensitive measurements of blood constituents in asample using IR spectroscopy. In an operational block 610, the method600 comprises providing an absorbance spectrum of the sample. In anoperational block 620, the method 600 further comprises shifting theabsorbance spectrum to zero at an isosbestic wavelength. Water and aprotein within the sample have approximately equivalent absorptions atthe isosbestic wavelength.

[0195] Various embodiments of the present invention have been describedabove. Although this invention has been described with reference tothese specific embodiments, the descriptions are intended to beillustrative of the invention and are not intended to be limiting.Various modifications and applications may occur to those skilled in theart without departing from the true spirit and scope of the invention asdefined in the appended claims.

What is claimed is:
 1. A method for using spectroscopy to determine ananalyte concentration in a sample, the method comprising: producing anabsorbance spectrum of the sample; shifting the absorbance spectrum tozero in a wavelength region; and subtracting a water or other substancecontribution from the absorbance spectrum.
 2. The method of claim 1,wherein the wavelength region is at or near an isosbestic wavelength. 3.The method of claim 1, wherein the sample is a biological sample and thesubstance is a whole blood protein.
 4. The method of claim 1, whereinthe analyte concentration determination is insensitive to pathlength. 5.The method of claim 1, wherein subtracting the water contribution fromthe absorbance spectrum comprises: providing a reference waterabsorbance spectrum; scaling the reference water absorbance spectrum bymultiplying the reference water absorbance by a scaling factor, whereinthe scaled reference water absorbance spectrum approximately equals theabsorbance spectrum at a predetermined wavelength; and subtracting thescaled reference water absorbance spectrum from the absorbance spectrum.6. The method of claim 1, wherein subtracting the substance contributionfrom the absorbance spectrum comprises: providing a reference substanceabsorbance spectrum; scaling the reference substance absorbance spectrumby multiplying the reference substance absorbance by a scaling factor,wherein the scaled reference substance absorbance spectrum approximatelyequals the absorbance spectrum at a predetermined wavelength; andsubtracting the scaled reference water absorbance spectrum from theabsorbance spectrum.
 7. The method of claim 1, comprising subtracting aresidual interactive component contribution from the absorbancespectrum.
 8. The method of claim 7, wherein the residual interactivecomponent comprises components of a boundary layer between water andprotein.
 9. The method of claim 1, further comprising subtracting apotentially interfering substance contribution from the absorbancespectrum.
 10. The method of claim 9, wherein the potentially interferingsubstance comprises urea.
 11. The method of claim 1, further comprisingsubtracting a glucose contribution from the absorbance spectrum anddetermining a residual substance concentration in the sample.
 12. Themethod of claim 11, wherein the residual substance comprises lactate.13. The method of claim 11, wherein the residual substance comprisesurea.
 14. The method of claim 1, wherein the OD spectrum is fitted withreference glucose spectral data, thereby yielding a measurement ofglucose concentration in the sample.
 15. The method of claim 14, whereinthe absorbance spectrum is fitted with reference glucose spectral dataat at least one glucose absorbance maximum.
 16. The method of claim 15,wherein the at least one glucose absorbance maximum is at approximately9.25 μm.
 17. The method of claim 15, wherein the at least one glucoseabsorbance maximum is at approximately 9.65 μm.
 18. A method ofproviding pathlength insensitive measurements of blood constituents in asample using infrared (IR) spectroscopy, the method comprising:providing an absorbance spectrum of the sample; shifting the absorbancespectrum to zero at an isosbestic wavelength, wherein water and aprotein within the sample have approximately equivalent absorptions atthe isosbestic wavelength.